Transglutaminases are critically important enzymes in the assembly of the keratinocyte cornified envelope, as well as, structures of the hair follicle. It is clear that a particulate form of transglutaminase, TGK, is present in the skin and in the inner and outer root sheaths of the hair follicles; however, other enzymes possessing transglutaminase activity have also been identified in these structures. Because of this, distinguishing the contribution of each specific enzyme has been difficult. In addition, although transglutaminase function has been studied in cell culture models, these models do not faithfully reproduce in vivo differentiation. Transgenic TGk knockout mice would provide a unique system in which to study the function of TGk and to determine whether any of the known envelope precursors are selectively utilized by this enzyme in vivo. The availability of such mice would make it possible to study the effects of TGk deficiency on epidermal differentiation and disease and to assess its potential role during mouse development. The major goal of this Pilot and Feasibility Study is to develop TGk knockout mice. Our specific aims are (1) to construct a plasmid that will permit us to inactivate the mouse TGk gene in mouse embryonic stem (ES) cells and (2) to establish TGk deficient ES cell lines and TGk deficient chimeric mice. To accomplish this goal, we will transfect cultured mouse embryonic stem (ES) cells with the TGk DNA- containing targeting vector, isolate TGk-deficient clonal cell lines and use these to generate chimeric and TGk-knockout mice.